DESCRIPTION (Investigator's Abstract): The long-term goal of this research is to understand from a cell biological perspective how neurons establish and maintain distinct axonal and dendritic domains that differ in structure and function. This feature of nerve cells-referred to as neuronal polarity- is essential to normal neural function. Disruption of neuronal polarity is thought to contribute to the pathophysiology of many neurologic diseases. The aims for the coming award period are to elucidate the cellular mechanisms underlying the targeting and transport of membrane proteins and to assess the role that these processes play in the development of neuronal polarity. In the previous award period it was shown that basolateral targeting signals that contain a tyrosine motif target exogenous proteins to dendrites, where exogenous proteins carrying motifs in the cytoplasmic domain of endogenous dendritic protein mediate their targeting and will identify the sequence within an endogenous axonal protein that govern its polarization to the axon. The earliest event in the establishment of neuronal polarity occurs when one of several, initially identical processes undergoes a prolonged period of growth, becoming the cell's axon. In order to assess possible causal relationships between specification of the axon and the development of protein targeting, changes in the distribution of representative axonal and dendritic proteins labeled with Green Fluorescent Protein will be followed as cells begin axonal outgrowth. Further experiments will perturb the localization of an endogenous cell adhesion molecule involved in the signaling that governs the specification of polarity in order to assess its impact on axonal outgrowth and the development of polarity. After the initial sorting event, which is presumed to direct different proteins to distinct vesicle populations, transport vesicles must be targeted to appropriate domains within the cell. The expression of appropriately tagged axonal, dendritic, and uniformly distributed proteins will be used to define the populations of transport vesicles present in hippocampal neurons and to visualize their transport in living cells.